The Tumor Necrosis Factor Super Family (TNFSF) consists of approximately 20 cellular surface signaling proteins with a conserved trimeric structure. All members of the TNFSF perform key roles in the activation or control of the immune system. The main hurdle preventing clinical use of these proteins as adjuvants or direct therapies in diseases such as cancer has been the structural requirements for effective signaling.
CD40L is an example of a TNFSF family member. In its native signaling environment CD40L engages CD40 on the surface of an adjacent cell to initiate signaling. CD40 initially adopts a monomeric structure on the cell surface, which is then trimerized by three CD40 subunits binding the three binding sites of the CD40L trimer. This in turn allows the cytosolic region of CD40 to engage the trimeric TRAF receptors, which serve as signal mediators for many of the TNFSF members. In addition to the trimeric structure of the TNFSF members, robust signaling is also enabled by multiple CD40L-CD40 contacts adjacent to each other forming a cluster. This naturally occurs when two cells engage each other but does not occur if soluble CD40L is administered.
Currently therapeutic TNFSF protein (e.g., CD40L) administration is limited by the high dose needed to elicit a beneficial effect, which leads to toxic off target effects. Antibodies against TNFSF proteins, such as anti-CD40 antibodies, have been constructed to act as high affinity substitutes that bypass the need for polyvalent high avidity contacts. However these antibodies cannot crosslink in the correct trimeric structure for signaling unless the concentration of antibody is significantly high.
Therefore, there is a desire for products and methods for the display of trimeric proteins.